人可溶性CD26（SCD26）ELISA試劑盒說明書
FOR RESEARCH USE ONLY
Assay range：0.5pg/ml-35pg/ml 96 determinations
Purpose
This kit allows for the determination of SCD26 concentrations in Human serum, cell culture supernates and
other biological fluids
Principle of the assay
The kit assay Human SCD26 level in the sample, use
Purified Human SCD26 antibody to coat microtiter plate wells,
make solid-phase antibody, then add SCD26 to wells, Combined
SCD26 antibody which With HRP labeled, become antibody -
antigen - enzyme-antibody complex, after washing Compley,
Add TMB substrate solution,TMB substrate becomes blue color At
HRP enzyme-catalyzed, reaction is terminated by the addition of
a sulphuric acid solution and the color change is measured
spectrophotometrically at a wavelength of 450 nm. The
concentration of Human SCD26 in the samples is then
determined by comparing the O.D. of the samples to the standard
curve.
Materials provided with the kit
1 wash solution
20ml×1bott
le
7 Stop Solution
6ml×1
bottle
2
HRP-Conjugate
reagent
6ml×1
bottle
8
Standard
（32pg/ml）
0.5ml×1
bottle
3
Microelisa
stripplate
12well×8str
ips
9
Standard
diluent
1.5ml×1bot
tle
4 Sample diluent
6ml×1
bottle
1
0
Instruction 1
5
Chromogen
Solution A
6ml×1
bottle
1
1
Closure plate
membrane
2
6
Chromogen
Solution B
6ml×1
bottle
1
2
Sealed bags 1
Specimen requirements
1. extract as soon as possible after Specimen collection,and
according to the relevant literature, and should be experiment
as soon as possible after the extraction. If it can’t, specimen can
be kept in -20 ℃ to preserve, Avoid repeated freeze-thaw
cycles.
2. Can’t detect the sample which contain NaN3, because NaN3
inhibits HRP active.
人可溶性CD26（SCD26）ELISA試劑盒說明書Assay procedure
1. Dilute and add sample:Dilute Original density Standard as follow
table:
16pg/ml
5 Standard 150µl Original density Standard+150µl Standard
diluent
8pg/ml
4 Standard 150µl 5 Standard+150µl Standard diluent
4pg/ml
3 Standard 150µl 4 Standard+150µl Standard diluent
2pg/ml
2 Standard 150µl 3 Standard +150µl Standard diluent
1pg/ml
1 Standard 150µl 2 Standard +150µl Standard diluent
2. Add sample: Set blank wells separay (blank comparison
wells don’t add sample and HRP-Conjugate reagent, other each
step operation is same). testing sample well. add standard 50µl
in the Microelisa stripplate accuray,add Sample dilution
40µl to testing sample well, then add testing sample 10µl
(sample final dilution is 5-fold), add sample to wells , don’t
touch the well wall as far as possible, and Gently mix.
3. Incubate: After closing plate with Closure plate
membrane ,incubate for 30 min at 37℃.
4. Configurate liquid: 30-fold(or 20-fold) wash solution diluted
30-fold (or 20-fold) with distilled water and reserve.
5. Washing: Uncover Closure plate membrane, discard Liquid,
dry by swing, add washing buffer to every well, still for 30s then
drain, repeat 5 times, dry by pat.
6. Add enzyme: Add HRP-Conjugate reagent 50µl to each well,
except blank well.
7. Incubate: Operation with 3.
8. Washing: Operation with 5.
9. Color:Add Chromogen Solution A 50ul and Chromogen
Solution B 50ul to each well, evade the light preservation for
10 min at 37℃
10.Stop the reaction:Add Stop Solution50µl to each well, Stop the
reaction(the blue color change to yellow color).
11. Assay:take blank well as zero , Read absorbance at 450nm
after Adding Stop Solution and within 15min.
Steps description
Standard, Sample
diluent
Add Standard, Sample diluent, incubate for 30 min at
37℃.
Wash 5 time,Add HRP-Conjugate reagent, incubate for 30 min at
37℃.
Wash 5 times,Add Chromogen Solution A and B, incubate for 10 min
at 37℃.
Add Stop Solution
Read absorbance at 450nm within 15 min
calculate
Calculate
Take the standard density as the horizontal, the OD value
for the vertical ,draw the standard curve on graph paper, Find out
the corresponding density according to the sample OD value by
the Sample curve, multiplied by the dilution multiple, or
calculate the straight line regression equation of the standard
curve with the standard density and the OD value ,with the
sample OD value in the equation, calculate the sample density,
multiplied by the dilution factor, the result is the sample actual
density.
人可溶性CD26（SCD26）ELISA試劑盒說明書Important notes
1. The kit takes out from the refrigeration environment should be
balanced 15-30 minutes in the room temperature, ELISA
plates coated if has not use up after opened, the plate should be
stored in Sealed bag.
2. washing buffer will Crystallization separation, it can be
heated the water helps dissolve when dilute . Washing does not
affect the result.
3. add Sample with sampler Each step, And proofread its
accuracy frequently, avoids the experimental error. add
sample within 5 min, if the number of sample is much ,
recommend to use Volley .
4. if the testing material content is excessively higher (The
sample OD is bigger than the first standard well ),please dilute
Sample (n-fold), Please diluente and multiplied by the
dilution factor.（×n×5）.
5. Closure plate membrane only limits the disposable use, to
avoid cross-contamination.
6. The substrate evade the light preservation.
7. Please according to use instruction strictly, The test result
determination must take the microtiter plate reader as a
standard.
8. All samples, washing buffer and each kind of reject should
according to infective material process.
9. Do not mix reagents with those from other lots.
Storage and validity
1．Storage： 2-8℃.
2．validity： six months
